| Inhibition of Alkaline Phosphatase from Pearl Oyster Pinctada fucata by o-Phthalaldehyde： Involvement of Lysine and Histidine Residues at the Active Site |
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| 引用本文： | 陈洪涛,谢莉萍,于振艳,张荣庆.Inhibition of Alkaline Phosphatase from Pearl Oyster Pinctada fucata by o-Phthalaldehyde： Involvement of Lysine and Histidine Residues at the Active Site[J].清华大学学报,2005,10(4):414-420. |
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| 作者姓名： | 陈洪涛 谢莉萍 于振艳 张荣庆 |
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| 作者单位： | [1]Institute of Marine Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China [2]Protein Science Laboratory of the Ministry of Education, Tsinghua University, Beijing 100084, China |
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| 基金项目： | Supported by the National High-Tech Research and Development（863） Program of China （No. 2003AA603430） |
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| 摘 要： | Alkaline phosphatase from Pinctada fucata was inactivated by o-phthalaldehyde （OPA）. The inactivation followed pseudo first-order kinetics with a second rate constant of 0.167 （mmol/L）^-1·min^-1 at pH 7.5 and 25℃. A Tsou＇s plot analysis showed that inactivation occurred upon formation of one isoindole group. The OPA-modified enzyme lost the ability to bind with the specific affinity column and the presence of substrates or competitive inhibitors protected the enzyme from inactivation. The results revealed that the OPA-reaction site was at the enzyme substrate binding site. Prior modification of the enzyme by lysine or histidine specific reagent abolished formation of the isoindole derivatives, suggesting that lysine and histidine residues were involved in the OPA-induced inactivation. Taken together, OPA inactivated the alkaline phosphatase from Pinctada fucata by cross-linking lysine and histidine residues at the active site and formed an isoindole group at the substrate binding site of the enzyme.
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| 关 键 词： | 碱性磷酸酶 苯二醛 化学特性 动态性质 |
| 收稿时间： | 2004-05-14 |
| 修稿时间： | 2004-05-14 |
Inhibition of Alkaline Phosphatase from Pearl Oyster Pinctada fucata by o-Phthalaldehyde: Involvement of Lysine and Histidine Residues at the Active Site |
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| Hongtao Chen, &#x;ÚÛ, Liping Xie, &#x;
, Zhenyan Yu, î&#x;,Rongqing Zhang, &#x;.Inhibition of Alkaline Phosphatase from Pearl Oyster Pinctada fucata by o-Phthalaldehyde: Involvement of Lysine and Histidine Residues at the Active Site[J].Tsinghua Science and Technology,2005,10(4):414-420. |
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| Authors: | Hongtao Chen &#x;ÚÛ Liping Xie &#x;
Zhenyan Yu î&#x; Rongqing Zhang &#x; |
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| Institution: | aInstitute of Marine Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China;bProtein Science Laboratory of the Ministry of Education, Tsinghua University, Beijing 100084, China |
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| Abstract: | Alkaline phosphatase from Pinctada fucata was inactivated by o-phthalaldehyde (OPA). The inactivation followed pseudo first-order kinetics with a second rate constant of 0.167 (mmol/L)?1·min?1 at pH 7.5 and 25°C. A Tsou's plot analysis showed that inactivation occurred upon formation of one isoindole group. The OPA-modified enzyme lost the ability to bind with the specific affinity column and the presence of substrates or competitive inhibitors protected the enzyme from inactivation. The results revealed that the OPA-reaction site was at the enzyme substrate binding site. Prior modification of the enzyme by lysine or histidine specific reagent abolished formation of the isoindole derivatives, suggesting that lysine and histidine residues were involved in the OPA-induced inactivation. Taken together, OPA inactivated the alkaline phosphatase from Pinctada fucata by cross-linking lysine and histidine residues at the active site and formed an isoindole group at the substrate binding site of the enzyme. |
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| Keywords: | alkaline phosphatase Pinctada fucata chemical modification kinetics |
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